Cancer develops by the accumulation of numerous mutations that usurp signaling, proliferation, and apoptotic pathways. Specifically, the p53 pathway is disrupted in the majority of cancers. Multiple mechanisms inactivate this pathway including mutations of the p53 gene itself and overexpression of the MDM2 and MDM4 genes which encode inhibitors of p53. Using mouse models, we have shown the functional importance of Mdm2 and Mdm4, an Mdm2-related protein, as inhibitors of p53 activity. Deletion of Mdm2 or Mdm4 in mice causes an embryo lethal phenotype that is completely rescued by concomitant deletion of p53. Additionally, mice heterozygous for Mdm2 or Mdm4 show increased p53 activity in response to ionizing radiation suggesting that even two fold differences in p53 inhibitors may affect tumor onset. Such two fold changes in MDM2 or MDM4 levels may affect tumor incidence in humans as an MDM2 polymorphism that increases MDM2 levels is associated with earlier age of onset of cancer. In response to DNA damage, the Mdm2 gene is also alternatively spliced to generate small Mdm2 isoforms that inhibit Mdm2 and thus activate p53. The role of these isoforms in tumorigenesis is controversial. We will test the hypothesis that alterations in the levels of the p53 inhibitors Mdm2 and Mdm4 affect the timing of tumor onset and the kinds of tumors that develop. First, we plan to examine the effects of Mdm2 or Mdm4 haploinsufficiency in tumor prone mice with p53 or K-ras mutations. We will generate a mouse model that cannot make the Mdm2 isoforms to examine effects on tumorigenesis. We will also characterize transgenic mice that overexpress Mdm4 to examine its role in tumorigenesis. Lastly, we will examine the relationship of Mdm2 and Mdm4 in inhibition of p53 by replacing the Mdm2 gene with the Mdm4 gene.